Intracellular ATP concentrations can vary 10-fold or more depending upon a cell's state of health or developmental stage. It is of great value to be able to measure fluctuations in intracellular ATP levels as a means of investigating e.g. the effects of drugs, toxins, hormones, environmental agents or disease on cells.
There is apparently at present no convenient method for analysing the concentration of ATP in vivo. For instance, in Dementieva et al (1996) Biochemistry (Moscow) Vol 61, No. 7., the intracellular concentration of ATP was measured in E. coli by calculating the total amount of ATP present using a recombinant luciferase, and dividing by an estimated total cell volume.
Such an indirect approach can at best produce only an estimate of the actual ATP concentration.
The measurement of ATP concentration in cells has also been performed using an in vitro coupled assay, such as that disclosed in the Sigma Diagnostic Kit Catalog No. 366, in which Phosphoglycerate kinase is used to convert 3-phosphoglycerate to 1,3 diphosphoglycerate in an [ATP]-dependent fashion. The 1,3 diphosphoglycerate is then converted to glyceraldehyde-3-P concommitantly with conversion of NADH to NAD, which can be monitored spectroscopically. The assay has a dynamic range up to 1 mM; the expected range is 380-620 .mu.m when used with blood cells.
However it can be seen that, as with all coupled assays, the test is inevitably cumbersome to perform. Additionally it could not readily be adapted for in vivo use. It would thus be a contribution to the art to provide materials and methods which overcome some of the drawbacks of the prior art.